This proposal represents a close collaborative effort between the Jardetzky and Longnecker laboratories to focus our understanding of Epstein-Barr virus (EBV) entry into target cells on potential therapeutic intervention. The entry of EBV into both epithelial cells and B cells is directly relevant to the Program Announcement for AIDS-related malignancies. Both cell types are EBV target tissues in the human host and the presence of EBV has been linked to cancerous growth in both tissues, in particular after the development of AIDS. The development of inhibitors of EBV entry into B cells and epithelial cells may prevent virus spread to susceptible cells inAIDS patients - while the linkage between EBVandAIDS-associated cancers is strong, not all EBV infected tissues become cancerous in AIDS patients, suggesting that infection of the right tissue at the right time may be important to disease development. We anticipate that a better understanding of EBV entry will be important for the development of therapeutics to treat EBV associated disease in AIDS patients and we are directly pursuing this goal by validating targets for inhibitor development, using both structural and functional approaches. Discoveries related to the proposed work will also be relevant to a general understanding of herpesvirus entry and replication and provide new tools to analyze key steps in herpesvirus penetration of cells. We feel that developing the tools to predict and validate protein:protein interaction inhibitors will become of increasing importance for both drug discovery and novel chemical biology approaches directed towards understanding fundamental mechanisms in biology. Our overall hypothesis is that there are multiple steps in the process of EBV entry into cells that are good targets for inhibitor development, such as the interactions of the EBV envelope glycoproteins that are critical to activating viral and cellular membrane fusion. The focus of this proposal is to develop lead inhibitors for known interaction sites in the EBV gp42 and gH/gL proteins that will block membrane fusion in B cells and to establish targets for inhibitor development in the gH/gL complex as a means to inhibiting both B cell and epithelial cell entry. Small molecule and peptide inhibitors of EBV entry will establish a basis for further development of antiviral drugs for the treatment of EBV infection and related diseases. The proposed combination of biophysical and structural methods with cell-based infection and celhcell fusion assays provides a comprehensive approach to the identification of therapeutic targets and lead compounds for the treatment of EBV-associated malignancies in patients with AIDS.